Abstract Intermediate filaments (IFs), microfilaments and microtubules are the three major cytoskeletal protein families found in most mammalian cells. Of these three families, IFs are the largest (>70 genes) and mutations in IF genes cause or predispose to more than 70 common and orphan human diseases. In digestive-organ epithelia, keratin polypeptides 8, 18, 19, and 20 (K8/K18/K19/K20) are the major IF proteins; and mutations in K8/K18/K19 predispose their carriers to acute or chronic liver disease progression. Disruptions of normal IF cytoplasmic organization is a defining feature of most IF diseases. A critical current challenge is the absence of targeted therapies for any IF disease, and the lack of selective IF-stabilizing compounds; in contrast to well-established compounds such as paclitaxel that stabilize microtubules. An important biological property of many IFs, including K18/K19, is that they serve as major caspase substrates during apoptosis. Furthermore, disease-causing keratin mutations that involve the caspase recognition motif interfere with caspase digestion. K18 is cleaved sequentially at two aspartates (one conserved in most IFs and the second K18-specific), while K19 is cleaved only at the conserved aspartate. However, the significance of any keratin proteolysis during apoptosis is unknown, except for our recent results that suggest an important role in allowing K18 filament reorganization and protecting cells from necrosis. Our proposal tests two hypotheses: First, caspase digestion of keratins is critical to permitting keratin filament reorganization and to protecting cells from injury; Second, small-molecule compounds can be identified that alleviate cytoskeletal structural alterations caused by keratin mutations and, thereby, provide potential novel treatment options for cytoplasmic keratin diseases. We already have extensive preliminary results to support both hypotheses. We propose to test the two hypotheses using four specific aims: (i) Determine the function of K18 proteolysis in digestive organs during apoptosis; (ii) Define the function of K19 proteolysis during apoptosis in the intestine; (iii) Identify and characterize small-molecule compounds that stabilize disassembled mutant keratins; (iv) Test the therapeutic utility and selectivity of compounds identified in Aim 3. The proposed approach combines state-of-the-art high throughput screening, molecular and cell biological approaches, and transgenic animal models to translate basic science discoveries into new fundamental knowledge and potential treatments. Our findings should help establish the functional role of keratin proteolysis by caspases during apoptosis, and clarify the pathogenesis of some disease-causing keratin mutations that interfere with caspase digestion. We anticipate being able to define novel compounds, or reposition current FDA-approved drugs, for normalizing mutation-induced keratin disorganization. These drugs may serve as potential therapies for IF diseases or as powerful research tools for studying fundamental cell biologic principles of the mammalian IF cytoskeleton.